A number of eukaryotic expression vectors have been developed for use as DNA vaccines. They showed varying abilities to initiate immune responses;however, there is little data to indicate which of these vectors will b...A number of eukaryotic expression vectors have been developed for use as DNA vaccines. They showed varying abilities to initiate immune responses;however, there is little data to indicate which of these vectors will be the most useful and practical for DNA vaccines in different species. This report examines the use of five expression vectors with different promoters and Kozak sequence to express the same hemagglutinin (HA) protein of an H6N2 avian influenza virus for DNA vaccination in chickens. Although intramuscular vaccination with seven DNA constructs elicited no or limited measurable H6 HA antibody responses in Hy-Line chickens, variable reduction in virus shedding for either oropharyngeal or cloacal swabs post-virus challenge were observed. This indicated that all DNA constructs generated some levels of protective immunity against homologous virus challenge. Interestingly, lower dose (50 or 100 μg) of plasmid DNAs consistently induced better immune response than higher dose (300 or 500 μg). In the transfection experiments there appeared to be a hierarchy in the in vitro expression efficiency in the order of pCAG-optiHAk/ pCAG-HAk > pCI-HAk > VR-HA > pCI-HA > pCI-neo-HA > pVAX-HA. Since the level of in vitro expression correlates with the level of immune response in vivo, in vitro expression levels of the DNA constructs can be used as an indicator for pre-selection of plasmid vaccines prior to in vivo assessment. Moreover, our results suggested that the Kozak sequence could be used as an effective tool for DNA vaccine design.展开更多
AIM: To investigate the genetic constitution of an escape mutant H5N1 strain and to screen the presence of possible amino acid signatures that could differentiate it from other Egyptian H5N1 strains.METHODS: Phylogene...AIM: To investigate the genetic constitution of an escape mutant H5N1 strain and to screen the presence of possible amino acid signatures that could differentiate it from other Egyptian H5N1 strains.METHODS: Phylogenetic, evolutionary patterns and amino acid signatures of the genes of an escape mutant H5N1 influenza A virus isolated in Egypt on 2009 were analyzed using direct sequencing and multisequence alignments.RESULTS: All the genes of the escape mutant H5N1 strain showed a genetic pattern potentially related to Eurasian lineages. Evolution of phylogenetic trees of different viral genes revealed the absence of reassortment in the escape mutant strain while confirming close relatedness to other H5N1 Egyptian strains from human and avian species. A variety of amino acid substitutions were recorded in different proteins compared to the available Egyptian H5N1 strains. The strain displayed amino acid substitutions in different viral alleles similar to other Egyptian H5N1 strains without showing amino acid signatures that could differentiate the escape mutant from other Egyptian H5N1.CONCLUSION: The genetic characteristics of avian H5N1 in Egypt revealed evidence of a high possibility of inter-species transmission. No amino acid signatures were found to differentiate the escape mutant H5N1 strain from other Egyptian H5N1 strains.展开更多
The rapid epidemic of highly pathogenic A/H5N1 avian influenza virus by transmission from poultry to humans triggered global unrest in the pandemic of novel influenza. If a human trophic strain of avian influenza viru...The rapid epidemic of highly pathogenic A/H5N1 avian influenza virus by transmission from poultry to humans triggered global unrest in the pandemic of novel influenza. If a human trophic strain of avian influenza viruses replicates in livestock including pigs and chickens, it may have high infectivity and pathogenicity to humans. The most effective method of reducing the outbreaks of influenza would be prophylaxis with an effective vaccine as well as anti-viral drugs including Oseltamivir and Zanamivir hydrate. In this study, chicken antiserum against A/H5N1 virus was produced: the antisera from immunized adult chicken had a strong binding activity to A/H5N1 viral antigens by ELISA. Furthermore, the antiserum strongly inhibited hemaggregation of erythrocytes and cytopathic effects in MDCK cells, indicating a strong neutralization activity against A/H5N1 infections. Interestingly, the mortality rate of chicks inoculated with A/H5N1 virus was dramatically decreased with the antiserum injection. These results suggest that antiserum may be a potentially effective protective and therapeutic modality for A/H5N1 infection.展开更多
Avian influenza is the most contagious disease not only in poultry, but also in humans. Avian influenza in humans occurs mainly in Southeast Asia, but no human-to-human pandemic has occurred. Meanwhile, outbreaks of a...Avian influenza is the most contagious disease not only in poultry, but also in humans. Avian influenza in humans occurs mainly in Southeast Asia, but no human-to-human pandemic has occurred. Meanwhile, outbreaks of avian influenza in poultry occur on a global scale and cause a large economic loss. Migration antibodies passed from mother birds via eggs are said to be an important component of the immune system that protects birds from infection. Thus, the immunity status of mother birds can determine the ability of offspring to defend against infection. In this study, we investigated the presence of anti-avian influenza virus antibody in chickens hatched on a poultry farm in Indonesia and examined the involvement of migratory antibodies in protecting against virus infection by infectious experiments of highly pathogenic avian influenza in chickens. Blood was collected from randomly selected chicks, and antibodies against avian influenza virus were evaluated in all birds. Since these young birds had no history of vaccination, the antibodies were deemed to have been transferred from the mother birds. The enzyme-linked immunosorbent assay antibody titer in each bird varied. Infection of these birds with highly pathogenic avian influenza virus A/H5N1 intra-nasally resulted in a high mortality rate in chicks with low antibody titers but a low mortality rate in chicks with high antibody titers. These findings indicate that migratory antibody prevented highly pathogenic avian influenza A/H5N1 infection in chicks, suggesting that such a preventive effect could also be expected with outdoor natural infection.展开更多
The rapid spread of the highly pathogenic A/H5N1 avian influenza virus among domestic birds and its transmission to humans has induced world-wide fears of a new influenza pandemic. A/H5N1 has infected over 300 people ...The rapid spread of the highly pathogenic A/H5N1 avian influenza virus among domestic birds and its transmission to humans has induced world-wide fears of a new influenza pandemic. A/H5N1 has infected over 300 people since 1997, and has shown a mortality rate of over 50%. The high mortality in human cases is thought to be enhanced by the excessive secretion of various endogenous factors, including cytokines and interleukins, stimulated by viral infections. Chickens infected with A/H5N1 viruses experience sudden death without showing severe clinical symptoms or inflammation. However, severe hemorrhage and congestion are seen in various tissues in sporadic chicken cases of A/H5N1-infections, especially in the pulmonary tissues, thus indicating that there is ischemia due to vascular abnormalities. Our previous studies have focused on the expression pattern of endothelin-1, which modulates the vascular tone via endothelin receptors. An Indonesian sporadic strain of A/H5N1 virus was intranasally administered to 10-day-old chicks, and the expression of endothelin was examined in the infected birds. All birds died within five days of inoculation, and had moderate inflammation accompanied by severe hemorrhage and congestion in the lungs. Immunohistochemical studies showed enhanced expression of endothelin-1 in the infected lungs. In addition, the real-time PCR analyses revealed that endothelin-1 and endothelin receptor A mRNA were significantly elevated in the birds with A/H5N1 infections. Subsequently, H5N1-infected birds were inoculated with bosentan hydrate, a competitive antagonist of endothelin receptors. Interestingly, the mortality rate of the infected birds was dramatically decreased in a dose-dependent manner by the administration of bosentan hydrate. The pathological lesions, including congestion and hemorrhage in the pulmonary tissues, were clearly inhibited. These findings are promising, and suggest that endothelin receptor antagonists are a potential treatment for the highly pathogenic avian flu.展开更多
文摘A number of eukaryotic expression vectors have been developed for use as DNA vaccines. They showed varying abilities to initiate immune responses;however, there is little data to indicate which of these vectors will be the most useful and practical for DNA vaccines in different species. This report examines the use of five expression vectors with different promoters and Kozak sequence to express the same hemagglutinin (HA) protein of an H6N2 avian influenza virus for DNA vaccination in chickens. Although intramuscular vaccination with seven DNA constructs elicited no or limited measurable H6 HA antibody responses in Hy-Line chickens, variable reduction in virus shedding for either oropharyngeal or cloacal swabs post-virus challenge were observed. This indicated that all DNA constructs generated some levels of protective immunity against homologous virus challenge. Interestingly, lower dose (50 or 100 μg) of plasmid DNAs consistently induced better immune response than higher dose (300 or 500 μg). In the transfection experiments there appeared to be a hierarchy in the in vitro expression efficiency in the order of pCAG-optiHAk/ pCAG-HAk > pCI-HAk > VR-HA > pCI-HA > pCI-neo-HA > pVAX-HA. Since the level of in vitro expression correlates with the level of immune response in vivo, in vitro expression levels of the DNA constructs can be used as an indicator for pre-selection of plasmid vaccines prior to in vivo assessment. Moreover, our results suggested that the Kozak sequence could be used as an effective tool for DNA vaccine design.
文摘AIM: To investigate the genetic constitution of an escape mutant H5N1 strain and to screen the presence of possible amino acid signatures that could differentiate it from other Egyptian H5N1 strains.METHODS: Phylogenetic, evolutionary patterns and amino acid signatures of the genes of an escape mutant H5N1 influenza A virus isolated in Egypt on 2009 were analyzed using direct sequencing and multisequence alignments.RESULTS: All the genes of the escape mutant H5N1 strain showed a genetic pattern potentially related to Eurasian lineages. Evolution of phylogenetic trees of different viral genes revealed the absence of reassortment in the escape mutant strain while confirming close relatedness to other H5N1 Egyptian strains from human and avian species. A variety of amino acid substitutions were recorded in different proteins compared to the available Egyptian H5N1 strains. The strain displayed amino acid substitutions in different viral alleles similar to other Egyptian H5N1 strains without showing amino acid signatures that could differentiate the escape mutant from other Egyptian H5N1.CONCLUSION: The genetic characteristics of avian H5N1 in Egypt revealed evidence of a high possibility of inter-species transmission. No amino acid signatures were found to differentiate the escape mutant H5N1 strain from other Egyptian H5N1 strains.
文摘The rapid epidemic of highly pathogenic A/H5N1 avian influenza virus by transmission from poultry to humans triggered global unrest in the pandemic of novel influenza. If a human trophic strain of avian influenza viruses replicates in livestock including pigs and chickens, it may have high infectivity and pathogenicity to humans. The most effective method of reducing the outbreaks of influenza would be prophylaxis with an effective vaccine as well as anti-viral drugs including Oseltamivir and Zanamivir hydrate. In this study, chicken antiserum against A/H5N1 virus was produced: the antisera from immunized adult chicken had a strong binding activity to A/H5N1 viral antigens by ELISA. Furthermore, the antiserum strongly inhibited hemaggregation of erythrocytes and cytopathic effects in MDCK cells, indicating a strong neutralization activity against A/H5N1 infections. Interestingly, the mortality rate of chicks inoculated with A/H5N1 virus was dramatically decreased with the antiserum injection. These results suggest that antiserum may be a potentially effective protective and therapeutic modality for A/H5N1 infection.
文摘Avian influenza is the most contagious disease not only in poultry, but also in humans. Avian influenza in humans occurs mainly in Southeast Asia, but no human-to-human pandemic has occurred. Meanwhile, outbreaks of avian influenza in poultry occur on a global scale and cause a large economic loss. Migration antibodies passed from mother birds via eggs are said to be an important component of the immune system that protects birds from infection. Thus, the immunity status of mother birds can determine the ability of offspring to defend against infection. In this study, we investigated the presence of anti-avian influenza virus antibody in chickens hatched on a poultry farm in Indonesia and examined the involvement of migratory antibodies in protecting against virus infection by infectious experiments of highly pathogenic avian influenza in chickens. Blood was collected from randomly selected chicks, and antibodies against avian influenza virus were evaluated in all birds. Since these young birds had no history of vaccination, the antibodies were deemed to have been transferred from the mother birds. The enzyme-linked immunosorbent assay antibody titer in each bird varied. Infection of these birds with highly pathogenic avian influenza virus A/H5N1 intra-nasally resulted in a high mortality rate in chicks with low antibody titers but a low mortality rate in chicks with high antibody titers. These findings indicate that migratory antibody prevented highly pathogenic avian influenza A/H5N1 infection in chicks, suggesting that such a preventive effect could also be expected with outdoor natural infection.
文摘The rapid spread of the highly pathogenic A/H5N1 avian influenza virus among domestic birds and its transmission to humans has induced world-wide fears of a new influenza pandemic. A/H5N1 has infected over 300 people since 1997, and has shown a mortality rate of over 50%. The high mortality in human cases is thought to be enhanced by the excessive secretion of various endogenous factors, including cytokines and interleukins, stimulated by viral infections. Chickens infected with A/H5N1 viruses experience sudden death without showing severe clinical symptoms or inflammation. However, severe hemorrhage and congestion are seen in various tissues in sporadic chicken cases of A/H5N1-infections, especially in the pulmonary tissues, thus indicating that there is ischemia due to vascular abnormalities. Our previous studies have focused on the expression pattern of endothelin-1, which modulates the vascular tone via endothelin receptors. An Indonesian sporadic strain of A/H5N1 virus was intranasally administered to 10-day-old chicks, and the expression of endothelin was examined in the infected birds. All birds died within five days of inoculation, and had moderate inflammation accompanied by severe hemorrhage and congestion in the lungs. Immunohistochemical studies showed enhanced expression of endothelin-1 in the infected lungs. In addition, the real-time PCR analyses revealed that endothelin-1 and endothelin receptor A mRNA were significantly elevated in the birds with A/H5N1 infections. Subsequently, H5N1-infected birds were inoculated with bosentan hydrate, a competitive antagonist of endothelin receptors. Interestingly, the mortality rate of the infected birds was dramatically decreased in a dose-dependent manner by the administration of bosentan hydrate. The pathological lesions, including congestion and hemorrhage in the pulmonary tissues, were clearly inhibited. These findings are promising, and suggest that endothelin receptor antagonists are a potential treatment for the highly pathogenic avian flu.
